Ovule wall with calcium oxalate in Asarum canadense
This is an ovule wall in the Gynoecium of Asarum canadense, or wild ginger. Calcium oxalate crystals are evident in the outer wall of the ovule tissue (ca. 2 um yellow features.) Imaging with crossed polarizers shows their birefringent crystalline nature, e.g., www.flickr.com/photos/14643312@N02/17921887815. These appear to be single crystals with typically one per cell. The safranin-o has stained nuclei and what I presume are lignin rich tissues, while the fast green has stained the cell walls.
The protocol was as follows. Specimens fixed in FAA (formaldehyde, acetic acid, ethanol) 48 hr. Dehydrated in 35, 50, 75, 85, 95, 99 % IPA in water, 6 hours each min. Infiltrated in xylene saturated with Paraplast for 2 days, followed by 2 changes of melted Praplast for 2 hours each. Embedded in Paraplast. Sectioned on a Spencer 820 microtome at 11 micron. Cleared in xylene, 5 min, 2X. Rehydrated in 99, 95, 80, 70 %IPA. Stained in Johansen's Safranin-O, Fast Green (24 hours in Safranin-O, 15 sec. in fast green). Cleared 2 X in xylene 10 min each and mounted with DPEX.
Photographed on a Spencer 42 petrographic polarizing microscope using an original magnification of 200X, using a Sony NEX 5N with a Leica MIKAS 1/3X adapter.
Ovule wall with calcium oxalate in Asarum canadense
This is an ovule wall in the Gynoecium of Asarum canadense, or wild ginger. Calcium oxalate crystals are evident in the outer wall of the ovule tissue (ca. 2 um yellow features.) Imaging with crossed polarizers shows their birefringent crystalline nature, e.g., www.flickr.com/photos/14643312@N02/17921887815. These appear to be single crystals with typically one per cell. The safranin-o has stained nuclei and what I presume are lignin rich tissues, while the fast green has stained the cell walls.
The protocol was as follows. Specimens fixed in FAA (formaldehyde, acetic acid, ethanol) 48 hr. Dehydrated in 35, 50, 75, 85, 95, 99 % IPA in water, 6 hours each min. Infiltrated in xylene saturated with Paraplast for 2 days, followed by 2 changes of melted Praplast for 2 hours each. Embedded in Paraplast. Sectioned on a Spencer 820 microtome at 11 micron. Cleared in xylene, 5 min, 2X. Rehydrated in 99, 95, 80, 70 %IPA. Stained in Johansen's Safranin-O, Fast Green (24 hours in Safranin-O, 15 sec. in fast green). Cleared 2 X in xylene 10 min each and mounted with DPEX.
Photographed on a Spencer 42 petrographic polarizing microscope using an original magnification of 200X, using a Sony NEX 5N with a Leica MIKAS 1/3X adapter.